Natural killer (NK) cells and lymphokine-activated killer (LAK) cells have been implicated in immunosurveillance against tumor cells and allograft rejection (T. Barlozzari, C. W. Reynolds, and R. B. Herberman, J. Immunol., 131, 1024, 1983; A. A. Rayner, E. A. Grimm, M. T. Lotze, E. W. Chu, and S. A. Rosenberg, Cancer, 55, 1327, 1985). These effector cells also play a role in the regulation of immune responses (R. B. Herberman and J. R. Ortaldo, Science, 214, 24, 1981) and the control of viral and bacterial infections (R. M. Natuk and R. M. Welsh, J. Immunol., 138, 877, 1987; Weinhold et al. Lancet, Apr. 23, 902-904 (1988)). Therefore, it is of potential importance to be able to efficiently prepare functional NK cells for adoptive transfer immunotherapy in humans.
It was shown that monocytes interfere with the activation of LAK activity by IL-2. L-leucine methyl ester (LME) and L-phenylalanine methyl ester (PME) were shown to remove monocytes from human peripheral blood mononuclear cells (PBMC). LME was also shown to deplete NK activity and NK cells (M. Hoyer, T. Meineke, W. Lewis, B. Zwilling, and J. Rinehart, Cancer Res., 46, 2834, 1986; D. L. Thiele and P. E. Lipsky, J. Immunol., 134, 786, 1985; Lipsky and Thiele, U.S. Pat. No. 4,752,602).
Monocytes have also been removed by their adherence to nylon-wool columns or by centrifugal elutriation in order to generate LAK cells at high cell density. However, these procedures for monocyte removal are tedious and complicated. Some LAK cell precursors may also adhere to the nylon-wool columns. Therefore, we have employed PME, at a concentration of about 1 to 5 mM, as a single step for monocyte depletion. We were able to generate LAK from PME-treated cells. We have shown that depletion of monocytes by PME allows generation of LAK cells by rIL-2 at a cell density of 5.times.10.sup.6 /mL or higher (Leung, K. H., Lymphokine Research, 6, Abstract #1718, 1987; European patent application 87107755.8, published Dec. 2, 1987, and U.S. Pat. No. 4,849,329.
Depletion of monocytes by PME, as with LME, also diminished the NK activity of the cells. However, whereas inhibition of NK activity by LME was irreversible, with PME, the NK activity is recovered within 18 hr by incubation of the cells in medium supplemented with FCS or IL-2 at 37.degree. C. This recovery process may not, however, be acceptable for infusion to a patient in an adoptive transfer therapy protocol. We have, therefore, studied the removal of monocytes using amino acid lower alkyl esters in the presence of other amino acid analogs to determine whether these analogs may protect NK cell activity from inhibition by the amino acid lower alkyl ester.